micromachines Article Assessment of Sub-Micron Particles by Exploiting Charge Differences with Dielectrophoresis Maria F. Romero-Creel, Eric Goodrich, Danielle V. Polniak and Blanca H. Lapizco-Encinas * ID Microscale Bioseparations Laboratory and Biomedical Engineering Department, Rochester Institute of Technology, Rochester, NY 14623, USA; [email protected] (M.F.R.-C.); [email protected] (E.G.); [email protected] (D.V.P.) * Correspondence: [email protected]; Tel.: +1-585-475-2773 Received: 7 July 2017; Accepted: 30 July 2017; Published: 2 August 2017 Abstract: The analysis, separation, and enrichment of submicron particles are critical steps in many applications, ranging from bio-sensing to disease diagnostics. Microfluidic electrokinetic techniques, such as dielectrophoresis (DEP) have proved to be excellent platforms for assessment of submicron particles. DEP is the motion of polarizable particles under the presence of a non-uniform electric field. In this work, the polarization and dielectrophoretic behavior of polystyrene particles with diameters ranging for 100 nm to 1 µm were studied employing microchannels for insulator based DEP (iDEP) and low frequency (<1000 Hz) AC and DC electric potentials. In particular, the effects of particle surface charge, in terms of magnitude and type of functionalization, were examined. It was found that the magnitude of particle surface charge has a significant impact on the polarization and dielectrophoretic response of the particles, allowing for successful particle assessment. Traditionally, charge differences are exploited employing electrophoretic techniques and particle separation is achieved by differential migration. The present study demonstrates that differences in the particle’s surface charge can also be exploited by means of iDEP; and that distinct types of nanoparticles can be identified by their polarization and dielectrophoretic behavior. These findings open the possibility for iDEP to be employed as a technique for the analysis of submicron biological particles, where subtle differences in surface charge could allow for rapid particle identification and separation. Keywords: dielectrophoresis; electrical double layer; electrokinetics; particle polarization 1. Introduction The separation and assessment of submicron particles are essential processes in chemical and biological analysis, with particular importance in the fields of nanotechnology and biotechnology. Microfluidic devices, due to their small size, are an ideal platform for the examination of submicron particles. Electrokinetic (EK) methods have proven to be one of the leading microfluidic techniques for the assessment, separation, and enrichment of nano- to micron-sized particles. Electric field driven techniques offer great flexibility, since a single stimulation force can be used to move both the particles and the suspending medium. Electroosmotic flow (EOF) is commonly used as a liquid and particle pumping mechanism due to the attractive advantage of requiring no mechanical parts, as this phenomenon exploits the electrical double layer (EDL) of the device substrate material [1]. However, EOF requires high voltages, which can lead to undesirable effects such as electrolysis and Joule heating [2–4]. Electrophoresis (EP) is another important EK mechanism that refers to the motion of charged particles relative to the suspending medium, i.e., EP exploits particles’ charge differences to enable particle separations; where distinct particles exhibit differential migration under the influence of an electric field. EP is a well-known phenomenon, used commonly as gel EP for the separation of proteins and DNA in many applications. Other variations of EP, such as capillary EP, isoelectric Micromachines 2017, 8, 239; doi:10.3390/mi8080239 www.mdpi.com/journal/micromachines Micromachines 2017, 8, 239 2 of 14 focusing, isotachophoresis, electrochromatography, and micellar electrokinetic chromatography, are also commonly used successful techniques for separating nano and micron-sized particles by exploiting electrical charge differences [5,6]. Dielectrophoresis (DEP) is a peculiar electric-field driven technique since it exploits particle polarization effects, not electrical charge, when particles are exposed to a non-uniform electric field [1]. Particles acquire a dipole moment under the effect of an electric field, the poles in this dipole moment are subjected to several forces which results in a net dielectrophoretic force acting on the particle [7]. This method has received significant attention [8–15] due to its flexibility, as DEP can occur in AC and DC electric fields and it can be used to manipulate charged and non-charged particles. There is a plethora of reports available in the literature focused on the dielectrophoretic analysis of macromolecules, viruses, and other submicron particles [16–21]. DEP offers great potential for the separation and enrichment of particles by exploiting slight differences in particle polarization—which is dictated by particle characteristics such as shape, size, and dielectric properties. Depending on the sign of the dipole moment, particles move in distinct directions [7] and can exhibit positive or negative dielectrophoretic behavior. A particle that is more polarizable than the suspending medium will migrate towards the regions of higher electric field gradient under positive DEP; while particles with lower polarizability than the medium would migrate away from these regions due to negative DEP [22]. Particle polarization is a complex phenomenon; many groups have studied the polarization of submicron particles and its relation to particle dielectrophoretic behavior [7,21–30]. Differences in dielectric properties, such as surface charge, can be exploited to achieve effective particle separation. Green and Morgan [25] separated 93 nm diameter particles into two subpopulations by means of DEP with AC potentials at high frequencies (MHz range). In later studies [26,27], this group characterized the dielectrophoretic response of sub-micron particles as a function of electrolyte composition and conductivity, frequency of the applied potentials and particle size. In particular, they analyzed the effects of interfacial polarization mechanism and EDL polarization. The effect of surface functionalization has also been studied [28] demonstrating that a reduction on particle surface conductance leads to changes in the dielectrophoretic behavior of the particles. The DEP response of submicron particles under the influence of high frequency (>1000 Hz) electric potential is well documented in the literature by excellent studies [25–30]. However, the behavior of these particles at low frequencies (<1000 Hz) is still an unexplored area. Insulator-based DEP (iDEP), a dielectrophoretic mode where insulating structures are employed to create non-uniform electric fields [31,32]; usually employs low frequency AC and DC electric potentials to achieve particle manipulation [15,23,33–36]. In iDEP microsystems, particles are exposed to several forces simultaneously (EOF, EP, and DEP), and the resulting particle motion is the net migration caused by the combination of all present forces [37,38]. Particles can be “trapped” between the insulating structures if the applied electric field is high enough to produce DEP forces than can overcome all other present forces [31]; and particles will “stream” along the electric field lines when the generated DEP forces are just comparable to EOF and EP [32]. The interplay between these forces can be fine-tuned to achieve a desired dielectrophoretic process, from particle identification to the enrichment of low-abundant species [39]. The present work is focused on studying how differences in a particle’s electrical charge can be used to achieve dielectrophoretic separation of particles or detect changes in a particle’s surface composition, employing DC and low frequency (<1000 Hz) electric potentials. This low frequency range has not been fully explored in dielectrophoretic microsystems. This work is organized as follows: We first discuss the fundamentals of dielectrophoretic force and particle polarization and the acquisition of a dipole moment. We follow this discussion with mathematical predictions obtained with COMSOL Multiphysics® (Version 4.4, COMSOL Inc., Burlington, MA, USA) of the motion of submicron polystyrene particles under positive and negative DEP. Our experimental results demonstrate that is possible to identify and separate submicron particles with similar characteristics (same size, same Micromachines 2017, 8, 239 3 of 14 shape, and same substrate material) by means of iDEP by exploiting differences in surface charge. These findings open the possibility for iDEP to be employed as a technique for the analysis of submicron biological particles, where subtle differences in surface charge could allow for rapid particle identification and separation. 2. Theory 2.1. Dielectrophoretic Force A dielectric particle immersed in dielectric medium will polarize under the presence of an electric field. The effective dipole moment for a spherical particle is given by: ! ! 3 m = 4p#mrp fCM E (1) where #m is the medium permittivity, rp is the particle radius, and fCM is the Clausius-Mossotti factor (fCM). The dielectrophoretic force exerted on a spherical particle is derived from the dipole moment and is a function of particle size and its relative polarizability compared to that of the suspending medium: ! 3 2 F DEP = 2p#mrpRe( fCM)rE (2) where rE2 refers to the gradient of the electric field squared.